Neurobio-logy of multiple myeloma and its therapeutical use - results of the pilot study with a control arm.

BACKGROUND: Myeloma cells, occupying a bone marrow niche, are influenced not only by neighbouring stroma cells but also by signals from the axons of sympathetic nervous system. The nervous system is directly involved in the process of myeloma progression. Among other cancers, patients with myeloma suffer the most difficult distress generating intensive adrenergic signals, causing its further progression. There is a question arising from these facts regarding whether psychological interventions, modulating a function of the nervous system, can further improve outcomes of myeloma treatments. We focus on interactions between myeloma cells and the nervous system. PATIENTS AND METHODS: Twelve patients with monoclonal gamapathy of indetermined significance (MGUS) or myeloma have participated in this study; eight in the interventional arm with the intervention of forgiveness therapy and four in the control arm. The patients were in various phases of their treatment, from active observation to immuno-chemotherapy and autologous stem cell transplant. Two major types of parameters were measured during the intervention: parameters of the activity of the disease (MGUS or myeloma) and psycho-neuro-immunological parameters of the patient, such as psychological depression, anxiety, and anger by the validated test PROMIS), as well as activity of the autonomic nervous system by heart rate variability, and immune profile by flow cytometry of peripheral blood. RESULTS: Patients who completed the forgiveness intervention showed improvement of depression, anxiety, and anger measured by PROMIS above population average, significant expansion of physiological plasma cells CD138+38+ (P = 0.04), B memory lymphocytes CD27+ (P = 0.02), and dendritic plasmacytoid cells CD123+ (P = 0.03). Parameters of heart rate variability such as parasympatic nervous system (PNS) index, sympatic nervous system (SNS) index, stress index, standard deviation of NN intervals (SDNN) and root mean square of the successive differences (RMSSD) had improved in a majority of patients. CONCLUSION: An intervention centered on forgiveness therapy was able to improve distress, reduce adrenergic signals in the autonomic nervous system, and restore parameters of the immune profile of patients with plasma cell dyscrasia who suffered from chronic stress caused by repressed anger and unforgiveness. Integrative treatment of myeloma can improve the quality of life of patients and thus affect the efficiency of immuno-chemotherapy. New randomised trials are warranted to test the integrative treatment of myeloma that might be able to improve overall survival.

Cardiovagal regulation and transcutaneous pO2 in breast cancer patients - a pilot study.

Vagal activity in patients with metastatic or recurrent breast cancer can predict their survival and it can be altered by behavioral, pharmacological and surgical interventions. Tumor oxygenation is important in defining the cellular metabolic microenvironment of human malignancies, O2-depleted areas coincide with nutrient and energy deprivation and with a hostile metabolic microenvironment. In our work, we simultaneously measured two oxygen-sensitive parameters in breast cancer patients; blood oxygen saturation (SpO2) and trans-cutaneous O2 partial pressure (tcpO2) in breast tissue. Concurrently, 5-minute beat-to-beat heart rate recording was carried out in order to get heart rate variability (HRV) data from time-domain analyses, frequency-domain analyses and entropy and symbolic dynamic non-linear methods. We compared these parameters in patients newly diagnosed with breast cancer, in patients after therapy and in healthy controls. We found lower tcpO2 in patients with presence of malignant tumor compared to those post-treatment and/or without presence of malignancy. We also detected lower 2UV% (two unlike variations) and entropy in non-linear HRV analysis in all breast cancer patients and these parameters associated with parasympathetic activity did not return to the values comparable with healthy individuals after anti-cancer therapy, contrary to tcpO2. Our findings show that breast tissue tcpO2 can recover after the anti-cancer treatment, but complex heart rate control and cardio-vagal regulation remain impaired. This supports the idea that cancer patients and survivors might benefit from non-pharmacological interventions aimed at enhancing vagal activity, such as HRV biofeedback or Yoga.

Urinary fluorescence analysis in diagnosis of bladder cancer.

Early diagnosis of bladder cancer is crucial for improvement of cancer specific survival and recurrence rate. We analyzed the possible role of fluorescence urine analysis in bladder cancer diagnosis. The cohort consisted of 20 healthy controls, 40 patients with hematuria and 75 patients with hematuria and histologically proven bladder tumor. Synchronous fluores- cence spectra with a 70 nm wavelength difference were recorded for (1:1-1:128) urine dilutions. Concentration matrices of synchronous spectra (CMSS) were used to classify samples into tested groups. CMSS analysis allowed us to distinguish patients with tumor from patients with hematuria with a sensitivity 55% and specificity 74.7%. This is comparable to the sensitivity and specificity of other non-invasive tests like BTA stat and nmP-22 (Bladder check®). Lower fluorescence inten- sity of Imax 280 nm and ratio of 280 nm to 450 nm was found to be associated with the presence of tumor. We have found an association of decreased fluorescence with the stage of the disease. Our data suggest that CMSS urine analysis has a potential role in the non-invasive diagnostic tests for bladder cancer, but it cannot replace the current diagnostic algorithm yet.

Consequences of the natural retinoid/retinoid X receptor ligands action in human breast cancer MDA-MB-231 cell line: Focus on functional proteomics.

The main intention of this study was the investigation of impact of natural biologically active ligands of nuclear retinoid/retinoid X receptors (all-trans and 9-cis retinoic acid) on proteomic pattern in human estrogen receptor negative breast cancer cell line MDA-MB-231. For this purpose, proteomic strategies based on bottom-up method were applied. The total cell proteins were extracted utilizing a commercially Radio-Immunoprecipitation Assay (RIPA) buffer and separated on 2D sodium dodecyl sulfate polyacrylamide gel electrophoresis (2D SDS-PAGE). The proteins were subsequently digested in-gel by trypsin and their characterization was achieved by MALDI-TOF/TOF. By employing PDQuest™ software, we identified more than 50 proteins affected by retinoic acid isomers. For more information, 9 proteins which are associated with tumor process were selected. We determined that derivatives of retinoic acid led to significantly reduced level of proteins belonging to metabolic pathway (e.g. glyceraldehyde-3-phosphate dehydrogenase or pyruvate kinase 2) or to other cellular processes as apoptosis, regulation of transcription process or epithelial-mesenchymal transition (e.g. annexins, nucleoside diphosphate kinase B, vimentin). On the other hand all-trans retinoic acid treatment indicates up-regulated effect for heterogeneous nuclear ribonucleoprotein A2/B1.

Protein synthesis inhibitors of natural origin for CML therapy: semisynthetic homoharringtonine (Omacetaxine mepesuccinate).

Omacetaxine mepesuccinate is a drug approved in 2014 by FDA for the use in CML therapy in patients resistant to at least two thymidine kinase inhibitors (TKIs). It possesses unique mechanism of anticancer activity that is principally different from mechanism of activity of TKIs. Omacetaxine mepesuccinate inhibits protein translation through prevention of the initial elongation step of protein synthesis and its use benefits CML patients possessing the BCR-ABL oncogene. Because of the superior activity of Omacetaxine in patients who became resistant to therapy with TKIs, FDA decided on the accelerated approval of this drug taking its consideration not only its activity as such but also a favorable benefit-to-risk profile in patients included into clinical studies.

Acridin-3,6-dialkyldithiourea hydrochlorides as new photosensitizers for photodynamic therapy of mouse leukemia cells.

Acridin-3,6-dialkyldithiourea hydrochlorides (AcrDTUs) have been evaluated as a new group of photosensitizers (PSs) for photodynamic antitumor therapy (PDT). Mouse leukemia cells L1210 were used for testing of AcrDTUs as the new PSs. The irradiation (UV-A light (365 nm), 1.05 J/cm(2)) increased cytotoxicity of all derivatives against L1210 cells more than ten times. The highest photocytotoxicity was found for propyl-AcrDTU with IC50=0.48±0.03 µM after 48 h incubation. A generation of the superoxide radical anion upon UV-A irradiation of propyl-AcrDTU was confirmed by in situ photochemical EPR experiments. To explain a mechanism of photocytotoxic action of AcrDTUs, an intracellular distribution of propyl-AcrDTU has been studied. It was found that AcrDTU in non-irradiated cells was not present in their nucleus but in the lysosomes and partly in the mitochondria, and sequestration of propyl-AcrDTU was dependent on pH in lysosomes. After irradiation, the cell death was induced by oxidative damage of lysosomal and mitochondrial membranes. Concerning the cell cycle, flow cytometry after PDT with propyl-AcrDTU showed a significant increase of the cells in the subG0 phase. Observed signs of necrosis, apoptosis, and autophagy indicate that PDT/AcrDTU leads to multiple cell death types (caspase independent apoptosis, necrosis, and autophagy).

Differences in pteridine urinary levels in patients with malignant and benign ovarian tumors in comparison with healthy individuals.

Pteridines belong to a class of fluorescent metabolites that are excreted by humans in urine and their concentrations can reflect various pathophysiological states. We quantified the differences in urinary pteridine levels in patients with malignant and benign ovarian tumors and in healthy individuals. Urine samples were centrifuged and supernatants were oxidized by MnO2 before analysis. Levels of neopterin, biopterin, and pterin were assessed by fluorescence analysis of human urine after HPLC separation. We have revealed that the median neopterin levels were higher in urine samples from patients with malignant (0.226 µmol/mmol creatinine) and benign ovarian tumors (0.150 µmol/mmol creatinine) than in healthy subjects (0.056 µmol/mmol creatinine). The median neopterin levels of patients with malignant tumors were higher (1.5-times) than in patients with benign tumors. The median biopterin level in urine of patients with benign ovarian tumors (0.268 µmol/mmol creatinine) was found to be very close to the level in patients with malignant ovarian tumors (0.239 µmol/mmol creatinine), and both were higher than in healthy samples (0.096 µmol/mmol creatinine). The levels of urine pterin followed a pattern similar to neopterin levels for both ovarian tumors, but their concentrations were about three times lower than neopterin levels.

Fluorescence analysis of urine and its potential for ovarian cancer screening.

Early diagnosis of ovarian cancer could lead to decreased mortality. We assessed the possible use of urine autofluorescence analysis in its diagnostics and screening.We analysed urine from 42 healthy volunteers, 35 patients with benign, and 36 patients with malignant ovarian tumors. Synchronous fluorescence spectra with a 70 nm wavelength difference were recorded for (1:1 - 1:1024) urine dilutions. Concentration matrices of synchronous spectra (CMSS) were used to classify samples into tested groups.CMSS analysis allowed us to distinguish patients with malignant tumors from healthy ones with a high sensitivity (91.67 %) and specificity (100 %), a positive predictive value (PPV) 100 % and a negative predictive value (NPV) 93.33 %. However, discrimination between benign and malignant ovarian tumors was weaker, with sensitivity 86.11 %, specificity 77.14 %, PPV 79.49 % and NPV 84.38 %. Fluorescence intensity and the position of peaks at 330 and 360 nm were found to be associated with the grade and stage, suggesting that different fluorescent metabolites may prevail at different stages of the disease.CMSS analysis of urine provides an alternative for ovarian cancer screening method development and could be used as a diagnostic test to detect the recurrence of the disease after therapy.

Intracellular distribution of 3,6-bis(3-alkylguanidino)acridines determines their cytotoxicity.

UNLABELLED: Cytotoxicity of two derivatives of 3,6-bis(3-alkylguanidino)acridines (GNDAs; pentyl- and hexyl-GNDA) was determined against three cell lines: a murine immortalized fibroblast cell line NIH-3T3, a human ovarian carcinoma cell line A2780, and a human neuroblastoma cell line SH-SY5Y. We found out that these GNDAs were cytotoxic against A2780 and NIH-3T3 cells but they showed only a marginal cytotoxicity against neuroblastoma cells SH-SY5Y. To explain differences in cytotoxicity, intracellular distribution of GNDAs was monitored. GNDAs were accumulated in A2780 and NIH-3T3 cells in the nuclei (fluorescence microscopy). In contrast to these cell lines, in SH-SY5Y cells, GNDAs were localized outside of the nuclei, at the plasma membrane and surroundings, extending also to the cytosol. This distribution of GNDAs was confirmed by an ImageStream Flow Cytometer. Acetylcholinesterase (AChE) activity in the SH-SY5Y cells decreased upon incubation with GNDAs. Kinetic studies showed that GNDAs were able to inhibit AChE by the same mode as tacrine (9-amino-1,2,3,4-tetrahydroacridine), a known inhibitor of AChE. A low cytotocity of GNDAs against SH-SY5Y cells could be caused by their affinity to AChE (the enzyme is localized mainly at the plasma membrane). The interaction of GNDAs with AChE may affect their intracellular distribution and consequently the cytotoxicity. KEYWORDS: acetylcholinesterase, acridine, neuroblastoma cell line SH-SY5Y.

Proteomic analysis of changes in the protein composition of MCF-7 human breast cancer cells induced by all-trans retinoic acid, 9-cis retinoic acid, and their combination.

Retinoic acid (all-trans and 9-cis) isomers represent important therapeutic agents for many types of cancers, including human breast cancer. Changes in protein composition of the MCF-7 human breast cancer cells were induced by all-trans retinoic acid, 9-cis retinoic acid, and their combination and subsequently proteomic strategies based on bottom-up method were applied. Proposed approach was used for the analysis of proteins extracted from MCF-7 human breast cancer cell line utilizing a commercially manufactured kit RIPA and separated on two dimensional (2D) sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) after treatment with both retinoic acid isomers. We found significant differences in occurrence of proteins probably affecting the cell migration process in tumour cells. Heat shock protein 27, ribonucleoprotein SmD3, and cofilin-1 were significantly upregulated after treatment with combination of individual retinoic acid isomers. On the other hand, AP-5 complex subunit beta-1 shows the different response. Thus, the results might help to find the answer to important medical questions on (i) the identification of signaling pathways affected by retinoic acid isomers or (ii) how the observed proteomic pattern might reflect the effectiveness of retinoic acids treatment.

Realgar (As4S4) nanoparticles and arsenic trioxide (As2O3) induced autophagy and apoptosis in human melanoma cells in vitro.

The aim of the present study was to compare the effect of realgar nanoparticles and arsenic trioxide (ATO) on viability, DNA damage, proliferation, autophagy and apoptosis in the human melanoma cell lines BOWES and A375. The application of various flow cytometric methods for measurements cell viability, DNA cell cycle, mitochondrial potential, lysosomal activity, and intracellular content of glutathione was used. In addition, quantitative PCR, western blotting and multiplex bead array analyses were applied for evaluation of redox stress, autophagic flux, and cell signaling alterations.The results showed that realgar treatment of studied cells caused modulation of cell proliferation, induced a block in G2/M phase of the cell cycle and altered phosphorylation of IκB, Akt, ERK1/2, p38, and JNK kinases, as well as decreased mitochondrial membrane potential. Additionally, it appeared that induction of cell death by both realgar and ATO was dose-dependent, when lower (0.3 µM) dosage increased lysosomal activity and induced autophagy and higher (1.25 µM) concentration resulted in the appearance of apoptosis, while pan-caspase inhibitor attenuated more efficiently realgar- than ATO-induced cell death. Furthermore, low concentrations of ATO and realgar nanoparticles increased the content of intracellular glutathione and elevated γ-H2AX expression confirmed DNA damage preferentially at higher concentrations of both drugs used. Further analysis revealed slight differences in time-dependent phosphorylation pattern due to both realgar and ATO treatments, while significant differences were noticed between cell lines. In conclusion, realgar nanoparticles and ATO treatment induced dose-dependent activation of autophagy and apoptosis in both melanoma cell lines, when autophagy flux was determined at lower drug concentrations and the switch to apoptosis occurred at higher concentrations of both arsenic forms.

Novel 3,6-bis(imidazolidine)acridines as effective photosensitizers for photodynamic therapy.

The photoeffect of new proflavine derivatives with DNA-binding and antitumour activities, 3,6-bis((1-alkyl-5-oxo-imidazolidin-2-yliden)imino)acridine hydrochlorides (AcrDIMs), was studied to evaluate them as potential photosensitizers for photodynamic antitumor therapy. EPR measurements showed that superoxide radical anion and singlet oxygen were produced upon irradiation of AcrDIMs with UV-A light (>300nm) in the presence of molecular oxygen. This indicates that AcrDIMs may act as photosensitizers. The most active pentyl-AcrDIM and hexyl-AcrDIM displayed photocytotoxic effect toward the mouse lymphocytic leukemia cell line L1210 and human ovarian cancer cells A2780. Antitumor activity of pentyl-AcrDIM increased as high as about 12 times (72h incubation) after irradiation of A2780 cells (365nm, 1.05J/cm(2)). The photocytotoxicity seems to be associated with oxidative stress. Concerning the cell cycle, flow cytometry showed an arrest in the S-phase already 4h after irradiation. In a comet assay, no genotoxicity of AcrDIMs was found. Typical morphologic changes and formation of DNA-ladders indicated induction of apoptotic cell death, though no activation of caspase-3 was observed. Investigation of intracellular localization of pentyl-AcrDIM confirmed its partial accumulation in mitochondria and lysosomes. After irradiation of the A2780 cells, colocalization of pentyl-AcrDIM with monodansylcadaverine, a lysosomal dye, was proven, suggesting that lysosomes in the irradiated cells may be involved in the cell death.

Subcellular localization of proflavine derivative and induction of oxidative stress--in vitro studies.

Acridines have been studied for several decades because of their numerous biological effects, especially anticancer activity. Recently, cytotoxicity of novel acridine derivatives, 3,6-bis((1-alkyl-5-oxo-imidazolidin-2-yliden)imino)acridine hydrochlorides (AcrDIMs), was confirmed for leukemic cell lines [Bioorg. Med. Chem.2011, 19, 1790]. The mechanism of action of the most cytotoxic hexyl-AcrDIM was studied in this paper focusing attention on a subcellular distribution of the drug. Accumulation of hexyl-AcrDIM in mitochondria was confirmed after labeling mitochondria with MitoRED using ImageStream Imaging Flow Cytometer. The derivative significantly decreased intracellular ATP level (reduction of ATP level was decreased by vitamin E), and induced oxidative stress (ROS production detected by DHE assay) as well as cell cycle arrest in the S-phase (flow cytometry analysis) already after short-time incubation and induction of apoptosis. Cytotoxicity of hexyl-AcrDIM is closely connected with induction of oxidative stress in cells.

Proteasome inhibition leads to altered signaling in the proteome of cisplatin-resistant human ovarian carcinoma cell line.

To address a precise view into molecular mechanisms of apoptotic signaling pathways after single- or combinatory treatments with specific NF-κB- or proteasome inhibitors and/or cisplatin (CDDP), flow cytometry and western blotting of the cell proteome in human ovarian chemosensitive- and CDDP-resistant cell lines were used. We report here that proteasome inhibition (but not NF-κB inhibition) caused marked alterations in the cell proliferation and cell cycle, as well as in the levels of signaling anti- and pro-apoptotic proteins PARP, NF-κB, IκB-α, Bcl-2, Bax, and lysosome-associated LAMP-1 and ATP-7B molecules in particular proteome fractions. These findings refer to the possibility of regulation of CDDP resistance, inclusive the capacity of lysosomes to export CDDP in certain human ovarian cancer cells by proteasome inhibition.

Fluorescence characteristics of human urine from normal individuals and ovarian cancer patients.

Early diagnostics of ovarian cancer is difficult, because there are no symptoms until the disease has progressed to an advanced stage. As urine contains many intrinsic fluorophores, modern fluorescence techniques are perspective candidates for new routine urine tests. The presented work deals with differences in the fluorescence of metabolites in urine of ovarian cancer patients comparing to healthy volunteers using the fluorescence excitation-emission matrices. The most serious differences were found in undiluted urine at the fluorescence emission wavelengths from 400 nm to 460 nm when excited at 310 - 390 nm. Statistical analyses of our data have shown a 5-fold reduction in the intensity of the peak at 330/420 nm (excitation/emission wavelength) for undiluted urine samples excreted by cancer patients as compared to those of normal donors. Moreover, the ratio of intensities of the peaks at 370/440 nm and at 330/420 nm is 18-times elevated in urine excreted by patients with ovarian cancer as compared to healthy urine samples. The observed changes could be interpreted as reduction of the presence of pyridoxic acid, whereas blue-fluorescing pteridines becomes dominant in excitation-emission matrices of cancer urine samples in comparison to healthy donors. We suggest pteridines, which are related to cellular metabolism, as suitable candidates for neoplasia-associated fluorescent markers in human urine. Our work showed that monitoring of human urine fluorescent metabolites offers an alternative for ovarian cancer screening.

DNA binding acridine-thiazolidinone agents affecting intracellular glutathione.

Three new acridine-thiazolidinone derivatives (2a-2c) have been synthesized and their interactions with calf thymus DNA and a number of cell lines (leukemic cells HL-60 and L1210 and human epithelial ovarian cancer cell lines A2780) were studied. The compounds 2a-2c possessed high affinity to calf thymus DNA and their binding constants determined by spectrofluorimetry were in the range of 1.37 × 10(6)-5.89 × 10(6) M(-1). All of the tested derivatives displayed strong cytotoxic activity in vitro, the highest activity in cytotoxic tests was found for 2c with IC(50) = 1.3 ± 0.2 µM (HL-60), 3.1 ± 0.4 µM (L1210), and 7.7 ± 0.5 µM (A2780) after 72 h incubation. The cancer cells accumulated acridine derivatives very fast and the changes of the glutathione level were confirmed. The compounds inhibited proliferation of the cells and induced an arrest of the cell cycle and cell death. Their influence upon cells was associated with their reactivity towards thiols and DNA binding activity.

Trends in the lung cancer incidence and mortality in the Slovak and Czech Republics in the contexts of an international comparison

INTRODUCTION: Lung cancer represents the most frequent cause of cancer-related deaths in the industrialized countries. The aim of this study was to analyze the lung cancer incidence and mortality and the possible reasons for any differences discovered in two neighboring Central European countries-the Slovak Republic. METHODS: We used linear regression model when analyzing incidence and mortality; the trends are presented with corresponding 95% confidence intervals (CI) and p-value with null hypothesis being constant with time. RESULTS: Statistically significant increase of age-standardized incidence (0.707/100,000/year, 95% CI 0.107-1.307, p = 0,025) and mortality (1.339/100,000/year, 95% CI 1.050-1.629, p < 0.0001) of the lung cancer was revealed in males in the Slovak Republic (1980-1991). On the contrary, values of both indicators were stabilized in the Czech Republic. Since year 1991-2005 a statistically highly significant decrease of both incidence and mortality values was observed in males, which was greater in the Slovak Republic. Peak of the curve was not reached in women population, while incidence and mortality values have significantly continuous growth in both countries. CONCLUSIONS: According to the lung cancer incidence and mortality trends in both countries (in correlation with smoking prevalence) we consider the support of efforts to change the attitude towards smoking predominantly in women and younger generation to be the most accurate action to reduce these trends (AU)

Trends in the kidney cancer incidence and mortality in the Slovak and Czech Republics in 1980-2005 - in the context of an international comparison.

Incidence rates of kidney cancer vary in global standards more than tenfold, the highest worldwide rates are repeatedly registered in the Czech Republic (hereafter CR) and in other geographically neighbouring countries in Central Europe, including the Slovak Republic (hereafter SR). The aim of the study was to analyse the changes in time trends of incidence and mortality from kidney cancer in two geographically close countries (CR and SR) and to compare detected differences with the worldwide data. In spite of high rates of incidence and its global growth in analyses in 1980 - 2005, the character of its progress in time was changed in both countries. While in 1980 - 1994 the incidence of kidney cancer in males and females in both analysed countries increased significantly, after 1994 (to 2005) stagnation in males in SR and significant slowdown of its growth in males in CR were reported. In females in SR after 1994 significant slowdown of the incidence growth was reported and in CR there was even its non-significant fall. Mortality trend in both sexes in both countries in 1980-2005 was slower than the incidence. After 1994 (to 2005) in males in SR statistically non-significant slowdown of mortality growth was reported, in CR it was statistically significant fall of mortality rates. In women after 1994 (to 2005) statistically non-significant decrease was reported, in CR the decrease was significant. The increase of total incidence of the disease is not explained only by the growth of asymptomatic localized tumors due to high quality diagnostic methods, but it likely reflects actual growth of new cases of the disease. Assumed partial cause of the mortality stabilization and slowdown of the incidence growth after its previous culmination in 1994 is the decline of smoking and obesity prevalence in the last decades in men, although this fact does not reflect situation in women. More striking mortality decrease in CR in comparison with SR might be influenced by potentially more radical surgical therapy (rate of the amount of surgery within primary therapy according to the data in National Cancer Registry CR raises, in SR the data are not available) and by more significant increase of the disease rate in the clinical stages I and II in CR (in SR only short-time data are available).

Synthetic isothiocyanate indole-3-ethyl isothiocyanate (homoITC) enhances sensitivity of human ovarian carcinoma cell lines A2780 and A2780/CP to cisplatin.

Isothiocyanates (ITCs), popular chemopreventive agents present in cruciferous vegetables, prove growth-inhibiting and apoptosis-inducing activities in cancer cell lines in vitro. Our study presents a new synthetic ITC derivate indol-3-ethyl isothiocyanate (homoITC) as an effective modulator of cellular proliferation and inducer of apoptosis with potential utility as an anticancer drug or a sensitizer to routinely used chemotherapeutic agent cisplatin (cis-Pt).
We analyzed the growth inhibitory effects of homoITC in the human ovarian carcinoma cell line A2780 and its cisplatin-resistant variant A2780/CP using MTT-test and its apoptosis-inducing properties by flow cytometry and caspase 3 activation. Combination index (CI) values from Calcusyn software were used to characterize the interactions of homoITC and cis-Pt as synergistic (CI1). Significant synergistic effect in growth inhibition of homoITC (5 - 15 microM) and cis-Pt (2.5 - 10 microM) on A2780 parental cell line (CI from 0.42 to 0.85) was also observed on A2780/CP resistant subline (CI from 0.18 to 0.73) for 10-50 microM cis-Pt concentrations and the same concentrations of homoITC. Synergy in growth inhibition correlated with the potential of homoITC to stimulate apoptosis induced by cis-Pt.We conclude that homoITC may be worth of further studies assessing its value in the ovarian cancer treatment and elucidating mechanisms of its action.

Modulation of markers associated with aggressive phenotype in MDA-MB-231 breast carcinoma cells by sulforaphane.

Metastasis as a complex process involves loss of adhesion, migration, invasion and proliferation of cancer cells. Sulforaphane (SFN) is one of naturally occurring cancer chemopreventive isothiocyanates found in cruciferous vegetables, consumption of which has been associated with reduced risk of cancer. In this study, we describe effect of SFN on various aspects determining invasive behavior of MDA-MB-231 human breast carcinoma cells. We studied modulation of molecules associated with epithelial to mesenchymal transition (EMT), hypoxic marker CA IX and mitochondrially located peripheral benzodiazepine receptor (PBR) using flow cytometry, gene expression of matrix metalloproteinases MMP1, 3, 7, 9, 14, transcription factors POU5F1 and Twist1 mRNA by RT PCR, and cytokine production by multiplex bead assay. SFN downregulated PBR and vimentin expression in a dose dependent manner, but significantly affected neither HIF-1alpha, nor CA IX protein expression, nor VEGF and GLUT1 mRNA levels. Among studied MMPs, MMP7 and MMP14 mRNA were downregulated while no apparent effect on MMP1, MMP3 and MMP9 was observed. Further, we found significant down regulation of Twist1 and POU5F1, transcription factors that mediate EMT and the self-renewal of undifferentiated embryonic stem cells. SFN reduced also the production of pro-inflammatory cytokines IL-1beta, IL-6, TNF-alpha, IFN-gamma, immunomodulating cytokine IL-4 and growth factors involved in angiogenesis PDGF and VEGF. Our study shows that SFN efficacy is associated with the reversal of several biological characteristics connected with EMT or implicated in the matrix degradation and extracellular proteolysis, as well as with reduced production of pro-inflammatory cytokines and pro-angiogenic growth factors in MDA-MB-231 cells.